The fact that people can be repeatedly infected with malaria, and that prototype malaria vaccines have not conferred long-term protection, raises the possibility that immunological memory after malaria infection is poor. Our previous research, in malaria-exposed adults in Thailand, has shown that both antibody and memory B cell responses to malaria antigens were induced and were stably maintained over time in the absence of reinfection. Moreover, malaria-specific immediate effector interferon (IFN)-3 (24 hours ex vivo restimulation) and cultured IL-10 (6 day secretion into culture supernatant) T helper cell responses to malaria antigens were more prominent in subjects who had long-lived antibodies or memory B cells specific to malaria antigens. We also found that although IFN-3-producing effector memory T cells to malaria do not persist in the absence of regular boosting, very infrequent malaria infections are sufficient to generate central memory T cell responses. Nevertheless, not everyone generates a response to every antigen and responses to some antigens may be more stable in some people than others. We hypothesize that the inflammation associated with acute or repeated malaria infection results in the generation of short-lived effector T and B cells, whereas individuals experiencing only occasional infections can develop and maintain long-lived memory responses. The aim of this project is thus to investigate the phenotypic and functional characteristics of malaria-specific memory B and T cells in individuals living in Northern Thailand who have had differing levels of prior exposure to malaria infection (including individuals experiencing acute infection), and to assess the long-term durability of these naturally-induced responses. We will identify the phenotypes of malaria-specific memory B cells by using fluorochrome-conjugated malaria antigens to label B cells, counter stain with antibodies to surface markers and analyse by multiparameter flow cytometry. The functions of memory B cells based on antibody production will be analysed by limiting dilution assay. Immediate and long-term cultured cytokine production from peripheral blood T cells - after stimulation with malaria antigens in vitro - will be determined by intracellular cytokine staining and flow cytometry, and by multiplex bead-based assay, respectively. Transcription factors involved in development and survival of memory B cells will be analysed by polymerase chain reaction. We will track the fate of malaria-specific human B cells in the days, months and years after infection to determine the kinetics of the induction, maintenance and decay of malaria specific memory responses. By investigating the characteristics and durability of immune responses among individuals with differing levels of immunity to malaria, we will be able to describe the underlying molecular basis for the generation of long-lived immunity to malaria. This work is of clear relevance to development and evaluation of effective malaria vaccines which would be of major health benefit to the more than 3 billion people at risk of malaria infection, worldwide. PUBLIC HEALTH RELEVANCE: The proposed study aims to characterise the phenotype, function and longevity of the malaria antigen-specific B cells and helper T cells that develop in Thai adults with differing histories of previous malaria infection. The purpose of the work is to identify markers of long-lived humoral immune responses to malaria, a disease that still causes 300 to 500 million clinical cases and up to 1 million deaths annually worldwide. The results from this study will provide fundamental information addressing important issues for malaria vaccine development and evaluation.